Faculty of Life Science, Institute of Biochemistry, Leipzig University, Leipzig, Germany.
Protein Sci. 2023 May;32(5):e4637. doi: 10.1002/pro.4637.
Noncanonical amino acids (ncAAs) for photo- and chemical crosslinking are powerful biochemical tools for studying and manipulating interactions between proteins both in vitro and in intact cells. Since the first crosslinking ncAAs were genetically encoded about 20 years ago, the technology has now ripened beyond the proof-of-principle demonstrations and is contributing to the study of relevant biological questions in the frame of modern integrative approaches. Here, we provide an overview of available photo-activatable ncAAs for photo-crosslinking and electrophilic ncAAs for genetically encoded chemical crosslinking (GECX), with a major focus on the most recent entries such as ncAAs for SuFEx click chemistry and photo-activatable ncAAs for chemical crosslinking. We present recent examples of the application of genetically encoded crosslinkers to capture protein-protein interactions and identify interaction partners in live cells, to investigate molecular mechanisms of protein function, to stabilize protein complexes for structural studies, to derive structural information about protein complexes from the physiological cell environment, up to perspective applications of GECX-ncAAs for the development of covalent drugs.
非天然氨基酸（ncAAs）用于光交联和化学交联，是研究和操纵体外和完整细胞中蛋白质相互作用的强大生化工具。自大约 20 年前首次遗传编码交联 ncAAs 以来，该技术已经超越了原理验证演示阶段，并为现代综合方法框架内的相关生物学问题的研究做出了贡献。在这里，我们提供了可用的光活化 ncAAs 用于光交联和遗传编码化学交联（GECX）的亲电 ncAAs 的概述，主要关注最新的条目，如用于 SuFEx 点击化学的 ncAAs 和用于化学交联的光活化 ncAAs。我们介绍了最近将遗传编码交联剂应用于捕获蛋白质-蛋白质相互作用并鉴定活细胞中的相互作用伙伴、研究蛋白质功能的分子机制、稳定蛋白质复合物进行结构研究、从生理细胞环境中获取蛋白质复合物的结构信息，以及用于开发共价药物的 GECX-ncAAs 的潜在应用的例子。
